This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Polyunsaturated fatty acids are oxygenated by the dioxygenases of lipoxygenase (LOX) family of enzymes. Nonheme iron is responsible for highly stereoselective initial hydrogen abstraction from the fatty acid and subsequently non-tethered oxygen binds to the radical. Soybean LOX-1 (SBL1) is the best-studied model of lipoxygenase catalysis. The studies are supported by the availability of high resolution x-ray crystal structures. SBL1 helped to establish many fundamental concepts of lipoxygenase catalysis, in particular these are the high specificity of the reaction and formation of a chiral product;e.g. 5S-hydroperoxide from arachidonic acid. The nature of products led to a hypothesis of the substrate being capable of adopting a reversed orientation of binding in the lipoxygenase active site. This hypothesis is open to debate due to lack of LOX structure with bound substrate. A recent study suggested a head-first substrate orientation for the first oxygenation by SBL1. In the absence of the X-ray structure with bound substrate, modeling was a useful approach to understanding concepts developed by analysis of the stereochemistry of the lipoxygenase reaction. Since LOX binds spin-labeled fatty acids it is possible to directly measure distances between doxylstearic acid (DSA) and nitroxide spin label to establish the mode of substrate binding. Dr. Gaffney nursed the plan to carry out such study, ACERT has responded to subsequent service request by providing distance measurements.